Bottom Line:
Bacterial populations was enumerated by qPCR coupled to PMA treatment.L. lactis UL719 was able to survive and proliferate under simulated human colon, did not alter microbiota composition, but failed to inhibit C. difficile.While a single dose of 19 μmol/L (5× the MIC) was not sufficient to inhibit C. difficile, nisin at 76 μmol/L (20×the MIC) was effective at killing the pathogen.

ABSTRACTClostridium difficile is the most frequently identified enteric pathogen in patients with nosocomially acquired, antibiotic-associated diarrhea and pseudomembranous colitis. Although metronidazole and vancomycin were effective, an increasing number of treatment failures and recurrence of C. difficile infection are being reported. Use of probiotics, particularly metabolically active lactic acid bacteria, was recently proposed as an alternative for the medical community. The aim of this study was to assess a probiotic candidate, nisin Z-producer Lactococcus lactis UL719, competitivity and nisin (Nisaplin(®)) capacity to inhibit C. difficile in a model of human colon. Bacterial populations was enumerated by qPCR coupled to PMA treatment. L. lactis UL719 was able to survive and proliferate under simulated human colon, did not alter microbiota composition, but failed to inhibit C. difficile. While a single dose of 19 μmol/L (5× the MIC) was not sufficient to inhibit C. difficile, nisin at 76 μmol/L (20×the MIC) was effective at killing the pathogen. Nisin (at 76 μmol/L) caused some temporary changes in the microbiota with Gram-positive bacteria being the mostly affected. These results highlight the capacity of L. lactis UL719 to survive under simulated human colon and the efficacy of nisin as an alternative in the treatment of C. difficile infections.

Mentions:
The microbiota was challenged by 5× and 20× the MIC vs. C. difficile ATCC43255. Nisin at 5× the MIC did not alter the microbiota which remained stable (data not shown) although minor variations in the metabolite production profile (Table 4). At a nisin concentration of 20× the MIC, total microbiota significantly decreased by 0.7 log10 (p < 0.008), as shown in Figure 4. Gram-positive bacteria were affected by this higher amount of nisin, with Ruminococcaceae group being the mostly altered (-3.7 log10) after 24 h. In a lesser extent, a reduction of 1.5 log10, 1.3 log10, and 1 log10 were recorded for Lachnospiraceae group, Lactobacillaceae/Leuconostocaceae group and bifidobacteria, respectively. After 24 h of nisin administration, all bacterial populations recovered their initial counts except Ruminococcaceae group which dropped to its minimum counts. While acetate and butyrate significantly decreased (p < 0.05) from 76.24 and 32.13 mmol/L to 69.12 and 26.29 mmol/L, propionate production increased by 13% (Table 4). Besides, a nisin concentration of 5× did not inhibit C. difficile, which counts remained close to control (C. difficile alone) (Figure 3). Conversely, nisin at 20× was effective at inhibiting C. difficile with a significant reduction (p < 0.001) of 2.3 log10 at 1 h that lasted for 8 h (Figure 3). C. difficile was not detected after 24 h in this model (data not shown).

Mentions:
The microbiota was challenged by 5× and 20× the MIC vs. C. difficile ATCC43255. Nisin at 5× the MIC did not alter the microbiota which remained stable (data not shown) although minor variations in the metabolite production profile (Table 4). At a nisin concentration of 20× the MIC, total microbiota significantly decreased by 0.7 log10 (p < 0.008), as shown in Figure 4. Gram-positive bacteria were affected by this higher amount of nisin, with Ruminococcaceae group being the mostly altered (-3.7 log10) after 24 h. In a lesser extent, a reduction of 1.5 log10, 1.3 log10, and 1 log10 were recorded for Lachnospiraceae group, Lactobacillaceae/Leuconostocaceae group and bifidobacteria, respectively. After 24 h of nisin administration, all bacterial populations recovered their initial counts except Ruminococcaceae group which dropped to its minimum counts. While acetate and butyrate significantly decreased (p < 0.05) from 76.24 and 32.13 mmol/L to 69.12 and 26.29 mmol/L, propionate production increased by 13% (Table 4). Besides, a nisin concentration of 5× did not inhibit C. difficile, which counts remained close to control (C. difficile alone) (Figure 3). Conversely, nisin at 20× was effective at inhibiting C. difficile with a significant reduction (p < 0.001) of 2.3 log10 at 1 h that lasted for 8 h (Figure 3). C. difficile was not detected after 24 h in this model (data not shown).

Bottom Line:
Bacterial populations was enumerated by qPCR coupled to PMA treatment.L. lactis UL719 was able to survive and proliferate under simulated human colon, did not alter microbiota composition, but failed to inhibit C. difficile.While a single dose of 19 μmol/L (5× the MIC) was not sufficient to inhibit C. difficile, nisin at 76 μmol/L (20×the MIC) was effective at killing the pathogen.

ABSTRACTClostridium difficile is the most frequently identified enteric pathogen in patients with nosocomially acquired, antibiotic-associated diarrhea and pseudomembranous colitis. Although metronidazole and vancomycin were effective, an increasing number of treatment failures and recurrence of C. difficile infection are being reported. Use of probiotics, particularly metabolically active lactic acid bacteria, was recently proposed as an alternative for the medical community. The aim of this study was to assess a probiotic candidate, nisin Z-producer Lactococcus lactis UL719, competitivity and nisin (Nisaplin(®)) capacity to inhibit C. difficile in a model of human colon. Bacterial populations was enumerated by qPCR coupled to PMA treatment. L. lactis UL719 was able to survive and proliferate under simulated human colon, did not alter microbiota composition, but failed to inhibit C. difficile. While a single dose of 19 μmol/L (5× the MIC) was not sufficient to inhibit C. difficile, nisin at 76 μmol/L (20×the MIC) was effective at killing the pathogen. Nisin (at 76 μmol/L) caused some temporary changes in the microbiota with Gram-positive bacteria being the mostly affected. These results highlight the capacity of L. lactis UL719 to survive under simulated human colon and the efficacy of nisin as an alternative in the treatment of C. difficile infections.